A mannosyl transferase system specific for retinylphosphate as a glycosyl carrier in rat liver microsomal membranes was characterized. The system was active and specific in the absence of detergent, under conditions which preserved the intactness of the membrane. Arrhenius plots of mannosylretinylphosphate (MRP) synthesis revealed two slopes with a transition temperature at 12 degrees C. The biologically inactive analog perhydroretinylphosphate (pRP) formed MpRP but this reaction had a single slope. RP, but not pRP, was active in protein mannosylation. SDS-PAGE revealed major bands at 15, 30, 45, and 60 thouand daltons. The inactive analog pRP formed MpRP: this mannolipid was still available to the enzyme for reversal by excess GDP. On the contrary, the active compound MRP, once synthesized, was no longer available for reversal unless detergent was added along with GDP. It is concluded that RP acts as a direct carrier of mannose in the biosynthesis of specific glycoproteins and that saturation of the double bonds in the polyenic chain of vitamin A renders the molecule inactive in this process.